PAPER CHROMATOGRAPHY: Separation of Metal Ions

CHEMISTRY 161 – EXPERIMENT 2 PAPER CHROMATOGRAPHY: Separation of Metal Ions

INTRODUCTION Chromatography is a general name given to a family of separation techniques. Originally, the term was applied to the separation of colored substances (CHROMA-meaning color), however, this is no longer a limitation. The common feature of all chromatographic methods is the movement of one phase, called the MOBILE PHASE, across another, the STATIONARY PHASE. The extent to which separation of the components in a mixture is achieved depends upon the relative interaction of these components with both phases.

Chromatographic techniques are usually classified according to the nature of the stationary phase, which may be either a liquid or a solid. If the stationary phase is a solid, the technique is referred to as ADSORPTION chromatography; if it is a liquid, the technique is called PARTITION chromatography.

In "pure" adsorption chromatography separation depends primarily on the extent to which the components of a mixture, travelling in the mobile phase, are adsorbed onto the surface of the stationary phase. Thus a weekly adsorbed substance travels more rapidly across the stationary phase than a strongly adsorbed substance.

Although there are important adsorption effects which occur in paper chromatography, for simplicity we will consider it to be a partition system, where the stationary phase is water supported by the cellulose molecules of the paper, and the mobile phase is a solution of an organic solvent and water. The mixture to be separated is spotted on the paper which is then placed in a closed container containing the mobile phase. The latter is allowed to percolate through the fibers of the paper by capillary action in either an ascending or descending direction. As the mobile phase moves across the paper, it carries along the components of the mixture at different rates. When the mobile phase has moved a distance sufficient to achieve separation, the paper is removed and dried. Visualization of the separated components is enhanced by spraying the CHROMATOGRAM with a suitable reagent. Identification of the components is made by calculating a value called Rf, which is the ratio of the distance a given component moves relative to the distance the mobile phase or solvent front moves. distance component moves R  f distance solvent moves

Comparison with Rf values for known standards under the same experimental conditions is then made.

In this experiment paper chromatography is used to separate a mixture of metallic cations.

PROCEDURE: 1. Obtain six micro tubes and capillary applicators. Place four or five drops of the following solutions in separate micro tubes and label each tube accordingly.

TUBE SAMPLE LABEL 1 Fe(NO3)3 Fe3+ 2 Cu(NO3)2 Cu2+ 3 Ni(NO3)2 Ni2+ 2-1 4 Co(NO3)2 Co2+ 5 UNKNOWN UNKNOWN NO. _____ 6 UNKNOWN UNKNOWN NO. _____

2. Spend the next few minutes trying to standardize your spotting technique. Test the application procedure by dipping a separate applicator (avoid contamination) into each of the sample solutions and touching it to a piece of scrap chromatography paper. There is no need to spot either unknown here. The liquid from the applicator should form a spot about 5 mm in diameter. Label each spot and save this test paper to practice your spraying technique after your chromatogram is running.

3. Next, obtain a rectangular piece of chromatography paper about 20 cm long by 10 cm wide. Draw a light pencil line about 2 cm from the longer edge as shown in Figure 1. Fold the paper in half parallel to the 10 cm edge and then again into thirds so the creases are as shown in Figure 1 when you unfold the paper.

4. Lightly label each lane with a pencil as shown in Figure 1 and apply a 5 mm spot of the appropriate cation solution on the bottom line in each of the first four sectors of the paper (spot the Fe+3 lighter than the rest). Apply one unknown to each of the last two sectors (see Figure 1). Then apply the unknowns twice more to their corresponding sectors. The unknown is less concentrated than the known solutions, so this procedure will compensate for that deficiency.

Figure 1 5. Obtain about 15 mL of the mobile phase (also called eluting solution) from the reagent stock bottle in the fume hood. CAUTION: THE MOBILE PHASE IS A MIXTURE OF HYDROCHLORIC ACID AND 2– BUTANONE, AN ORGANIC SOLVENT. SKIN CONTACT AND INHALATION OF THESE IRRITANTS SHOULD BE AVOIDED.

2-2 6. Transfer the eluting solution to a 600 mL beaker and cover it immediately with a watch glass. Check to make sure that all the spots on the chromatography paper are dry. Refold the paper into the cylindrical hexagon (see Figure 1) using the creases you made to create the sectors. Carefully handling the paper by the top edges, place the paper into the mobile phase with the sample spots at the bottom. The spots should not come in direct contact with the solution and try not to allow the paper to come in contact with the sides of the beaker. Again cover the beaker with the watch glass. The mobile phase will gradually rise by capillary action, carrying along the different cations at different rates. After the process has gone on for a few minutes, you may be able to see colored spots on the paper corresponding to the migration of the different cations.

7. While the experiment is proceeding, you might want to test the action of the staining reagent on the test paper you spotted previously. The staining reagent is a solution containing potassium ferrocyanide and potassium ferricyanide and forms an insoluble colored precipitate with many cations. CAUTION: STAINS SKIN AND CLOTHING TOO! ALSO THE STAINING REAGENT IS TOXIC UPON INGESTION. BE SURE TO WASH HANDS BEFORE LEAVING LAB.

In the fume hood, spray the test paper evenly with the staining reagent, getting the paper moist but not really wet. Take the paper to a sink and rinse it with water to remove excess staining reagent. The colored spots will not wash out. Finally correlate the color of the spots with the cations you spotted.

8. When the eluting solution has risen to within about 1-2 cm of the top of the chromatography paper, remove the chromatogram from the beaker and place it on a paper towel. Quickly draw a pencil line along the solvent front which may not be a straight line. When the chromatogram is fairly dry, spray it with the staining reagent to make the positions of the cations more visible. Again rinse the chromatogram with water to remove excess staining reagent. When the paper dries, carefully circle each spot with a pencil and measure the distance in centimeters from the straight pencil line on which you applied the cation to the center of the spot. Then measure the distance from the pencil line to the solvent front. Calculate an Rf value for each cation and for each spot in the unknown lanes. From these Rf values and the color of the spots, determine which of the cations are present in your unknown solutions.

2-3 Name______

REPORT SHEET

PAPER CHROMATOGRAPHY: Separation of Metal Ions

A. Observed colors on test paper: Ion Before Staining After Staining Fe3+ ______Cu2+ ______Ni2+ ______Co2+ ______

B. Chromatographic results:

Distance Distance Color Unknown Solvent Ion Rf after Ions Moved Moved Staining Present Fe3+ ______Cu2+ ______Ni2+ ______Co2+ ______Unk#______

______Unk#______

Show a sample calculation for Fe3+ only.

2-4 POST-LAB QUESTIONS:

1. Assuming the solvent front moved 3 cm in 5 minutes, why shouldn't the experiment be stopped at that point instead of waiting 20 minutes for the front to move 8 cm? 2. What effect might the following have on your results:

a) The paper came in contact with the sides of the beaker during the experiment. b) One sector of the paper came in contact with another sector during the experiment. c) So much solvent was used that the starting line was actually immersed in the solvent.

3. In this experiment it takes about 5 microliters of a solution to produce the each spot. If the Cu(NO3)2 solution contains about 6 grams of Cu2+ per liter, how many grams of the Cu2+ ions are there in one spot?